Comparative distribution of VIP in the central nervous system of various species measured by a new radioimmunoassay

Vasoactive intestinal polypeptide (VIP) occurs in high concentrations throughout the gut and the nervous system. The presence of VIP has been shown in a number of species, mainly by immunohistochemistry. The aim of the present study was to develop a new, highly specific VIP radioimmunoassay to investigate the distribution of VIP in the central nervous system of various vertebrate and invertebrate species. Different areas of the brain and spinal cord were removed from rats, chickens, turtles, frogs and fishes. The cerebral ganglia and the ventral ganglionic chain were investigated in the earthworm. The tissue samples were processed for VIP radioimmunoassay. Our results show that the antiserum used in the radioimmunoassay turned to be C-terminal specific, without significant affinity to other members of the VIP peptide family. Detection limit of the assay was 0.1 fmol/ml. Highest concentrations were found in the turtle diencephalon, followed by other brain areas in the turtle and rat. All other brain areas in the examined species contained significant levels of VIP. Immunoreactivity was also shown in the cerebral and ventral ganglia of the earthworm. In summary, our results show comparative quantitative distribution in representative species of the phylogenetic line, using the same experimental conditions.     

Influence of pinealectomy on levels of PACAP and cAMP in the chicken brain

One of the recently found functions of pituitary adenylate cyclase activating polypeptide (PACAP) is the modulation of circadian rhythms. Widespread distribution of PACAP-containing neurons and receptors has been shown in the chicken. Recently, we have demonstrated that PACAP levels oscillate in a circadian manner in the chicken brain. Daily variation in PACAP levels might be influenced by several regulatory mechanisms. Among the structures that may regulate PACAP levels, one candidate is the pineal gland. Therefore, in the present study, we investigated the effect of pinealectomy on the levels of PACAP in the chicken brain. Animals were kept under 12:12-h light-dark schedule. Pinealectomy was performed at 3 weeks of age; sham-operated animals were used as controls. The animals were sacrificed at 15 and 24 h 1 week after pinealectomy. The brainstem and diencephalon were removed, and tissue samples were processed for PACAP and cAMP radioimmunoassay (RIA).PACAP and cAMP levels showed nighttime elevations in both the sham-operated and pinealectomized animals, except for the PACAP content in the diencephalon of pinealectomized chicken. PACAP levels of pinealectomized animals were significantly higher in the diencephalon and brainstem as compared to the control animals at both time-points. Levels of cAMP correlated well with levels of PACAP. The present results provide evidence that the pineal gland has an inhibitory impact on PACAP-neurons in the chicken brainstem and diencephalon.     

Shedding of the interleukin-6 (IL-6) receptor (gp80) determines the ability of IL-6 to induce gp130 phosphorylation in human osteoblasts

Human osteoblasts produce interleukin-6 (IL-6) and respond to IL-6 in the presence of soluble IL-6 receptor (sIL-6R), but the cell surface expression of IL-6R and the mechanism of sIL-6R production are largely unknown. Three different human osteoblast-like cell lines (MG-63, HOS, and SaOS-2) and bone marrow-derived primary human osteoblasts expressed both IL-6R and gp130 as determined by flow cytometry and immunoprecipitation. However, the membrane-bound IL-6R was nonfunctional, as significant tyrosine phosphorylation of gp130 did not occur in the presence of IL-6. Phorbol myristate acetate induced a dramatic increase of both IL-6R shedding (i.e. the production of sIL-6R) and IL-6 release in osteoblast cultures, but the cell surface expression of gp130 remained unchanged. IL-6 complexed with sIL-6R, either exogenously introduced or derived from the nonfunctional cell surface form by shedding, induced rapid tyrosine phosphorylation of gp130. This effect was inhibited by neutralizing antibodies to either sIL-6R or gp130, indicating that the gp130 activation was induced by IL-6/sIL-6R/gp130 interaction. Protein kinase C inhibitors blocked phorbol myristate acetate-induced and spontaneous shedding of IL-6R resulting in the absence of sIL-6R in the culture medium, which in turn also prevented the activation of gp130. In conclusion, human osteoblasts express cell surface IL-6R, which is unable to transmit IL-6-induced signals until it is shed into its soluble form. This unique mechanism provides the flexibility for osteoblasts to control their own responsiveness to IL-6 via the activation of an IL-6R sheddase, resulting in an immediate production of functionally active osteoblast-derived sIL-6R.     

To thermoconform or thermoregulate? An assessment of thermoregulation opportunities for the lizard<Emphasis Type="Italic"> Zootoca vivipara</Emphasis> in the subarctic

The general model of thermoregulation of ectotherms predicts that thermally challenging environments select for evolution of thermoconformity. Studies of reptilian thermoregulation at climatic extremes are rare and, in the subarctic zone, completely lacking. Thermal characteristics of the habitat of the lizard Zootoca vivipara were studied in the subarctic zone, at the northern margin of its distribution, where lizard density was already extremely low. We found that, during the activity period, the preferred body temperatures of Z. vivipara were not available for a thermoconformer, but available for 7 h for a thermoregulator in an average day. Therefore, thermoconformity is unbeneficial and accurate thermoregulation should be the appropriate strategy. We hypothesise that the extremely low lizard abundance at our subarctic study site is caused by the short activity season and the large daily temperature fluctuations, with night temperatures occasionally falling below zero even during the activity period.     

Glutamate decarboxylase activity in Trichoderma viride conidia and developing mycelia

Glutamic acid decarboxylase (GAD) activity was measured in homogenates of conidia and both submerged and aerial mycelia of Trichoderma viride. The GAD activity in conidia had a temperature optimum at 30 degrees C and a pH optimum at pH 4. GAD was stimulated by EDTA (2 mM) and was insensitive to treatment with calmodulin antagonists calmidazolium (10 microM) or phenothiazine neuroleptics (60 microM). Cyclosporin A (up to 300 microM) partially inhibited GAD in the homogenate, but not in the supernatant obtained after centrifuging the homogenate. Attempts to release GAD activity from the homogenate using high ionic strength, detergents, or urea failed. Freezing-thawing led to the partial increase of activity in the conidial homogenate. These results indicate that GAD is a membrane-bound enzyme. The highest specific activity of GAD was present in the mitochondrial/vacuolar organellar fraction. Germination of conidia in the submerged culture led to a temporary decrease in GAD activity. After prolonged cultivation, the activity displayed quasi-oscillatory changes. The stationary state was characterized by a high GAD activity. The presence of gamma-aminobutyric acid in the submerged mycelia was demonstrated. In surface culture in the dark, GAD activity increased in a monophasic manner until conidia formation. The illumination of dark-cultivated mycelia by a white-light pulse caused a dramatic increase in GAD activity. Light-induced changes were not observed in mutants with delayed onset of conidiation. In the dark or upon illumination by light pulse, the increase of GAD activity preceded the appearance of conidia. Thus, GAD activity in T. viride is closely associated with its developmental status and may represent a link between differentiation events and energy metabolism.     

Metabolic Changes in Rat Brain After Prolonged Ethanol Consumption Measured by 1H and 31P MRS Experiments

SUMMARY 1. In vivo ¹H and ³¹P magnetic resonance spectroscopy techniques were applied to reveal biochemical changes in the rat brain caused by prolonged ethanol consumption.2. Three models of ethanol intoxication were used.3. ¹H MRS showed a significant decrease in the concentration of myo-inositol in the brain of rats fed with 20% ethanol for 8 weeks. This change is consistent with perturbances in astrocytes. On the other hand, N-acetyl aspartate and choline content did not differ from controls.4. ³¹P MRS did not reveal any significant changes in the high-energy phosphates or intracellular free Mg²⁺ content in the brain of rats after 14 weeks of 20% ethanol drinking. The intracellular pH was diminished.5. By means of a ³¹P saturation transfer technique, a significant decrease was observed for the pseudo first-order rate constant k _for of the creatine kinase reaction in the brain of rats administered 30% ethanol for 3 weeks using a gastric tube.6. The ¹H MRS results may indicate that myo-inositol loss, reflecting a disorder in astrocytes, might be one of the first changes associated with alcoholism, which could be detected in the brain by means of in vivo ¹H MRS.7. The results from ³¹P MRS experiments suggest that alcoholism is associated with decreased brain energy metabolism.8. ³¹P saturation transfer, which provides insight into the turnover of high-energy phosphates, could be a more suitable technique for studying the brain energetics in chronic pathological states than conventional ³¹P MRS.     

The possible role of information technology in radiotherapy II. The development of a biological dose distribution model in the 3-dimensional treatment planning of brain tumours

OBJECTIVE: Developing a new medical software based on the utilisation of information technology required in 3-dimensional treatment planning and modern radiotherapy. METHODS: The physical dose distribution programs were converted into biological meaning with the insertion of biological equivalence equations based on LQ model. Biological dose distributions and biological dose-volume histograms were generated. The treatment plans of a brain tumour patient were investigated to determine the dose burdening of the normal central nervous system tissues. RESULTS: Employing 3D conformal method, the dose of the vital mid-line structures decreased significantly, which possesses a more meaningful biological importance. Different treatment plans and different fractionation regimens could be compared to each other by utilising this kind of biological model. CONCLUSION: By employing information technology we succeeded in establishing a theoretical biological dose distribution system that could be visualised. The advantages of 3D treatment planning proved unambiguous. In the future this method will probably be suitable to choose the best therapeutic regimens.     

13C and 1H Nuclear Magnetic Resonance Study of Glycogen Futile Cycling in Strains of the Genus Fibrobacter

We investigated the carbon metabolism of three strains of Fibrobacter succinogenes and one strain of Fibrobacter intestinalis. The four strains produced the same amounts of the metabolites succinate, acetate, and formate in approximately the same ratio (3.7/1/0.3). The four strains similarly stored glycogen during all growth phases, and the glycogen-to-protein ratio was close to 0.6 during the exponential growth phase. ¹³C nuclear magnetic resonance (NMR) analysis of [1-¹³C]glucose utilization by resting cells of the four strains revealed a reversal of glycolysis at the triose phosphate level and the same metabolic pathways. Glycogen futile cycling was demonstrated by ¹³C NMR by following the simultaneous metabolism of labeled [¹³C]glycogen and exogenous unlabeled glucose. The isotopic dilutions of the CH₂ of succinate and the CH₃ of acetate when the resting cells were metabolizing [1-¹³C]glucose and unlabeled glycogen were precisely quantified by using ¹³C-filtered spin-echo difference ¹H NMR spectroscopy. The measured isotopic dilutions were not the same for succinate and acetate; in the case of succinate, the dilutions reflected only the contribution of glycogen futile cycling, while in the case of acetate, another mechanism was also involved. Results obtained in complementary experiments are consistent with reversal of the succinate synthesis pathway. Our results indicated that for all of the strains, from 12 to 16% of the glucose entering the metabolic pathway originated from prestored glycogen. Although genetically diverse, the four Fibrobacter strains studied had very similar carbon metabolism characteristics.     

A 4-Year Study of a Natural Tick-Borne Encephalitis Virus Focus in Hungary, 2010–2013

EcoHealth - A tick-borne encephalitis virus focus was identified in a former goat pasture that had been associated with a milk-borne encephalitis outbreak in 2007. Ticks and rodents were sampled...